Extended Data Fig. 10: De novo design and experimental characterization of the influenza hemagglutinin (HA) binder.
From: Design of protein-binding proteins from the target structure alone
a, Structure comparison of the stem region of group 1 HA and group 2 HA. The stem regions of H1 HA (A/Puerto Rico/8/1934) (left, PDB code: 1RU7) and H3 HA (A/Hong Kong/1/1968) (right, PDB code: 4WE4) are shown in cartoon and colored in pale cyan and pale green respectively, the key residues in the stem region are shown as sticks. Three major differences make the H3 HA stem region a more challenging target for designing de novo protein binders: the H3 HA stem region contains more polar residues and is more hydrophilic. Residues in H1 HA that are hydrophobic residues or small polar residues while the corresponding residues are polar or larger polar residues are highlighted in dashed circles; Trp21 adopts different configurations in H1 HA and H3 HA, and the targeting groove in H3 HA is much shallower and less hydrophobic; the H3 HA is glycosylated at Asn38, and the carbohydrate side chains cover the hydrophobic groove and protect the HA stem region from binding by antibodies or designed binders. The insert shows a more extended view of the Asn38 glycosylation site on H3 HA. b, Binding of H3 binder to the H3 HA (A/Hong Kong/1/1968) N38D mutant (left) and H1 HA (A/Puerto Rico/8/1934) (right) with BLI. Two-fold serial dilutions were tested for each binder and the highest concentrations and the binder affinities are labeled. The gray color represents experimental data and orange color represents fit curves. c, The FI6v3 antibody competes with the binder for binding to the influenza A H1 hemagglutinin (left) and influenza A H3 hemagglutinin (right). Yeast cells displaying the H3 binder were incubated with 10 nM H1 or H3 in the presence or absence of 2 μM FI6v3 antibody, and hemagglutinin binding to cells (y axis) was monitored with flow cytometry.
